The present invention relates generally to the field of biochemical laboratory instrumentation for different applications of measuring properties of samples on e.g. microtitration plates and corresponding sample supports. More particularly the invention relates to the improved and more efficient instrumental features of equipment used as e.g. fluorometers, photometers and luminometers.
The routine work and also the research work in analytical biochemical laboratories and in clinical laboratories are often based on different tags or labels coupled on macromolecules under inspection. The typical labels used are different radioactive isotopes, enzymes, different fluorescent molecules and e.g. fluorescent chelates of rare earth metals.
The detection of enzyme labels can be performed by utilizing its natural biochemical function, i.e. to alter the physical properties of molecules. In enzyme immunoassays colourless substances are catalysed by enzyme to colourful substances or non-fluorescent substances to fluorescent substances.
The colourful substances are measured with an absorption, i.e. photometric measurement. In the photometric measurement the intensity of filtered and stabilized beam is first measured without any sample and then the sample inside one plate is measured. The absorbance i.e. the absorption values are then calculated.
The fluorescent measurement is generally used for measuring quantities of fluorescent label substance in a sample. The most photoluminescence labels are based on molecular photoluminescence process. In this process optical radiation is absorbed by the ground state of a molecule. Due to the absorption of energy the quantum molecule rises into higher excited state. After the fast vibrational relaxation the molecule returns back to its ground state and the excess energy is released as an optical quantum. Due to losses in this process the average absorbed energies are higher than the average emitted energies.
A further measurement method is chemiluminescence measurement where emission of a substance is measured from a sample without excitation by illumination. Thus any photoluminometer can also be used as a chemiluminometer.
The typical instruments in analytical chemical research laboratories are the different spectroscopic instruments. Many of them are utilizing optical region of electromagnetic spectrum. The two common type of instruments are the spectrophotometers and the spectrofluorometers. These instruments comprise usually one or two wavelength dispersion devices, like monochromators. The dispersion devices make them capable to perform photometric and fluorometric measurements throughout the optical spectrum.
FIG. 1 illustrates an advanced prior art optical analyser, especially the optical components and the different optical paths. The instrument has two illumination sources, a continuous wave lamp (cw-lamp) 112a and a pulse lamp 112b. The cw-lamp can be used for continuous wave photoluminescence excitation and for absorption measurements.
Infrared part of radiation from the cw-lamp 112a is absorbed by a filter 104, and after transiting a stray-light aperture plate 105, the optical radiation is collimated with a lens 115a through an interference filter 114a located in a filter wheel 114.
The light beam is focused with a lens 113a, similar to the lens 114a, into a light guide 118, which isolates the measuring head thermally and mechanically. It also shields the measuring unit for the stray light from the cw-lamp. The optical radiation from an output aperture plate 106 of a light guide 118 is collimated with a lens 107, similar to the lens 115a. The radiation beam is reflected by a beam-splitter mirror 141 inside a mirror block 140, and passed through a sample well 181 and through an entrance window 122 of a photometric detector unit 132.
The mirror block 140 is located on the upper side of the sample. Its function is to reflect the horizontal light beam from the selected lamp downwards to the sample and to reflect a portion of this beam by a mirror 143 into a reference photodiode 119, and also to allow the emission from the sample to travel upwards to the detector 132.
The emission unit comprises optical components, which are lenses 133, 135, a filter 134a in filter slide 134, a combined shutter and aperture slide 136 and a detector 132, such as a photo-multiplier. The detector 132 is used in the fast photon counting mode where the pulses from photo-multiplier anode are first amplified and then fed through a fast comparator 191 and gate 192 counter 193. The comparator rejects the pulses, which are lower than the pre adjusted reference level. The fast counting electronics is equipped with a gate in the front of the counter. This gate is used in overall timings of the measurements.
The pulse-lamp unit is used in time-resolved photoluminescence measurement for long-living photoluminescence emission. It comprises a second lamp 112b, lenses 115b, 113b, and optical filters 114b in a filter slide for wavelength isolation. When this second lamp is used the mirror 141 must be rotated by 90 degrees in order to reflect the radiation to the sample. This can be achieved by using different optical modules for the two lamps.
There are certain limitations related to the prior art technology. It is often required to make several measurements from same samples, e.g. measuring of two or more photoluminescence emissions, as well as absorption and chemiluminescence measurements may be required. With the prior art instruments it is necessary to make the different measurements successively, and it may be necessary to make changes in the optics of the instrument between the different measurements. Therefore performing such measurements from a large number of samples tends to take a very long measurement time with the prior art instruments, and the reliability of the measurement results is not optimal.
There are also instruments, which have two measurement heads; a top measurement head and a bottom measurement head. Such instruments are disclosed e.g. in documents U.S. Pat. No. 6,187,267 and U.S. Pat No. 5,933,232. With this kind of instrument it is possible to make measurements also from below the sample, so this kind of instrument is more versatile for performing different measurements. However, the prior art instruments are not capable of performing different measurements simultaneously, nor capable of performing dual emission measurements. Performing different measurements successively from a large number of samples tends to take a long time.
The object of the present invention is to provide an optical instrument for laboratory measurements, wherein the described disadvantages of the prior art are avoided or reduced. The object of the invention is therefore to achieve a measurement instrument with improved efficiency for performing measurements from samples.
The object of the invention is achieved by providing an optical measurement instrument where there is an interface for a changeable optical module, the interface being adapted for at least one excitation beam and at least two emission beams. The object is further achieved by a changeable optical module for a measurement instrument, the module comprising a preferably dichroic mirror for dividing an emission beam into two emission beams, and a preferably dichroic mirror for separating the optical paths of emission and excitation beams. The invention allows performing various types of measurements by changing an optical module. The change of module and related parameters can be performed automatically controlled by software. It is also possible to easily upgrade the instrument for new types of measurements by just providing the instrument with a new optical module and the related software.
An optical measurement instrument according to the invention for measuring samples, comprising an illumination source for forming an excitation beam, a first detector for detecting a first emission beam, an interface for a changeable optical module directing the excitation beam received from the illumination source into the sample and directing an emission beam received from the sample to the first detector, is characterized in that the interface further comprises means for receiving a second emission beam from a same optical module.
The invention also applies to a changeable optical module for an optical measurement instrument, the module comprising means for receiving an excitation signal from an illumination source and means for directing the excitation to a sample, means for receiving an emission beam from the sample and means for outputting the emission beam received from the sample to a detector, which is characterized in that the module further comprises means for separating the emission beam into a first emission beam and a second emission beam, and means for outputting the first emission beam for a first detector, and means for outputting the second emission beam for a second detector.
The invention also applies to a process for measurement of samples with an optical measurement instrument comprising means for providing excitation of a sample and means for measuring two emissions from the sample, the process comprising the phases of
selecting a measurement mode,
selecting a possible excitation filter,
selecting a first emission filter for a first detector,
selecting at least one optical module for guiding the excitation beam into the sample and for guiding the first emission into the first detector,
performing the optical measurement, which is characterized in that a process for measuring two emissions from the sample comprises the phases of
selecting a second emission filter for a second detector,
selecting one and same optical module for guiding the excitation beam into the sample, for dividing the emission beam into first emission beam and a second emission beam, for guiding a first emission beam into the first detector and for guiding a second emission beam into the second detector.
A method according to the invention for optical measurement of samples comprising the steps of:
forming an excitation beam,
directing the excitation beam to a sample with an optical module,
acquisition of an emission beam from the sample, is characterized in that the method further comprises the steps of:
dividing the emission beam into a first emission beam and a second emission beam within said optical module,
guiding the first emission beam to a first detector,
guiding the second emission beam to the second detector,
converting the emission beams into emission signals in said detectors, and
processing the signals for providing measurement results.
Some preferred embodiments are described in the dependent claims.
An important advantage of the invention relates to achieving high measurement efficiency. Measurements of two emissions can be made simultaneously, and the time needed for the measurement is thus halved. Further efficiency is achieved due to the minimal attenuation of the optical paths.
There are also other important advantages related to the idea of placing into a same changeable optical module the mirror for dividing the emission into two emission beams and the mirror for separating the optical paths of emission and excitation beams. This way one measurement head can be used for both one-emission measurement and for two-emission measurement in an optimal way. If a second emission is not measured with the same measurement head as the first emission, the optical module in use can be easily changed into a module, which does not include the mirror for the second emission beam. This way it is possible to have one emission measurement without unnecessary attenuation caused by the mirror.
A further advantage relates to the ability to offer optional functions in measuring equipment. Equipment with a measurement head for one emission measurement can be easily upgraded into equipment, which has a measurement head for one emission or two emission measurements. For the upgrade it is only necessary to provide the equipment with an optical module, which includes a mirror for the second emission, en providing the equipment with the second detector, if not readily available in the equipment. The basic version of the equipment preferably includes the required optics for guiding the second emission beam from the optical module to the second detector.
A further advantage relates to the possibility to have a filter combined with the mirror; different types of measurements can be optimized by selecting mirror that substantially transmits the wavelength of the first emission beam and substantially reflects the wavelength of the second emission beam. This way the attenuation of the emissions can be minimized, and there is less need for further filtering of the emission beams.
One further advantage of the present invention is related to the fact that two emissions can be measured without changing the connections of the optical fibres. This way the measurement modes can be changed by software without any need for manual work such as connecting and disconnecting optical cables.
The invention also allows the use of direct optical coupling in emission detection in the top measurement head of the equipment; attenuation caused by optical fibres is thus avoided.